Severe alcoholic liver disease (ALD) has a high morbidity and mortality. Recent studies demonstrated that probiotics reversed alcohol-induced hepatic steatosis and inflammation, and improved liver enzymes in animal models and in patients. Our laboratory showed that administration of a probiotic strain, Lactobacillus rhamnosus Gorbach-Goldin (LGG), significantly improved liver enzymes and histology in alcohol treated mice. These mice also had reduced intestinal permeability and plasma endotoxin levels. Gut-derived endotoxins normally penetrate the gut epithelium only in trace amounts due to tightly regulated intestinal barrier function. However, endotoxin leakiness is increased under pathological conditions, including alcohol abuse. We recently reported that chronic alcohol intake in mice caused a marked reduction in the intestinal expression of the master transcription factor, hypoxia inducible factor (HIF), and two HIF related proteins, cathelin-related antimicrobial peptide (CRAMP) and intestinal trefoil factor (ITF), in the intestine. ITF plays a critical role in the formation and stabilization of the intestinal mucus barrier, enhancement of rapid intestinal repair (restitution), and modulation of mucosal differentiation processes and immune responses, while CRAMP is important in blocking intestinal bacterial overgrowth/dysbiosis. In addition, our preliminary data demonstrate a significant decrease of short chain fatty acids (SCFAs), in particular, butyric acid, in the intestinal lumen following alcohol exposure in rodents. Our hypothesis is that alcohol consumption causes decreased intestinal CRAMP and ITF expression and intestinal lumen butyric acid concentration, resulting in bacterial overgrowth/dysbiosis and disrupted gut-barrier function; LGG supplementation attenuates these deleterious effects and protects the liver from alcohol-induced injury. To test this hypothesis, we will have 3 specific aims. Specific aim 1 will determine the role of CRAMP and the beneficial effects of LGG on CRAMP in alcohol-mediated changes in gut microbiota homeostasis. The experiments will be performed using a metagenomic approach in CRAMP knockout mice. Specific aim 2 will determine the role of LGG in promoting ITF expression and will evaluate whether ITF positively modulates intestinal tight junctions as a mechanism for improved intestinal barrier integrity. We will use goblet and intestinal epithelial cell culture, and recombinant ITF to delineate the effect of ITF. Specific aim 3 will determine the role and mechanisms of butyric acid in the beneficial effects of LGG on intestinal tight junctions using a butyric transporter knockout mouse model. This study will have a major impact on the development of probiotic-based new therapeutic strategy for the prevention and treatment of ALD.